Department of Oral Surgery, Ninth People's Hospital, College of Stomatology, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology & Shanghai Research Institute of Stomatology, Shanghai, People's Republic of China.
Department of Endodontics, Jinan Stomatological Hospital, Jinan, 250001, Shandong, People's Republic of China.
J Mol Histol. 2019 Oct;50(5):427-434. doi: 10.1007/s10735-019-09838-8. Epub 2019 Jul 3.
The accurately establishment of the eruptive pathway is of vital importance. The mechanisms governing tooth eruption pathway remain little known. This study is to elucidate the roles of Semaphorin 3A (Sema 3A) in mouse tooth eruptive pathway. C57BL/6 mice (11-13 and 15-17 days after birth) were chosen to observe eruptive pathway of mouse lower first molar. Expressions of Sema 3A and its receptor neuropilin 1 and plexin A1 were detected. Osteoclasts were identified by TRAP staining. Co-localization of Sema 3A and osteoclast maker CD68 was detected by double immunofluorescence staining. Picrosirius red staining was applied to observe collagen fibers during mucosal penetration phase. In vitro, Bone marrow-derived macrophages (BMMs) were prepared from 4 week C57BL/6 mice to observe the effect of Sema 3A on the differentiation of BMMs into osteoclasts by TRAP staining. Expressions of Sema 3A was observed by immunofluorescence and western blotting. At osseous eruption phase, many TRAP-positive multi-nucleated cells were distributed around occlusal alveolar bone. The positive expressions of Sema 3A were observed in the multi-nucleated cells. Fluorescence double staining showed that Sema 3A and CD68 were co-expressed in osteoclasts. Its receptor neuropilin 1 and plexin A1 were also found in osteoclasts. In vitro, Sema3A negatively regulated osteoclast differentiation. At mucosal penetration, occlusal alveolar bone had been completely resorbed and collagen fires were gradually degraded for eruptive pathway. Similar positive expressions of Sema 3A and its receptor neuropilin 1 and plexin A1 were also found in the mucosal penetration pathway. Sema 3A gets involved in the establishment of mouse tooth eruptive pathway by modulating osteoclast activity. Sema3A should be considered as a novel nervous agent or a potential biomarker for mouse tooth eruptive pathway.
准确建立萌出途径至关重要。控制牙齿萌出途径的机制知之甚少。本研究旨在阐明 Sema 3A（Semaphorin 3A）在小鼠牙齿萌出途径中的作用。选择 C57BL/6 小鼠（出生后 11-13 天和 15-17 天）观察小鼠下第一磨牙的萌出途径。检测 Sema 3A 及其受体神经纤毛蛋白 1 和神经丛蛋白 A1 的表达。通过 TRAP 染色鉴定破骨细胞。通过双重免疫荧光染色检测 Sema 3A 与破骨细胞标志物 CD68 的共定位。应用苦味酸红染色观察黏膜穿透期胶原纤维。在体外，从 4 周龄 C57BL/6 小鼠制备骨髓来源的巨噬细胞（BMMs），通过 TRAP 染色观察 Sema 3A 对 BMMs 向破骨细胞分化的影响。通过免疫荧光和 Western blot 观察 Sema 3A 的表达。在骨萌出阶段，许多 TRAP 阳性多核细胞分布在咬合牙槽骨周围。在多核细胞中观察到 Sema 3A 的阳性表达。荧光双重染色显示 Sema 3A 与 CD68 在破骨细胞中共同表达。其受体神经纤毛蛋白 1 和神经丛蛋白 A1 也在破骨细胞中发现。在体外，Sema3A 负调控破骨细胞分化。在黏膜穿透时，咬合牙槽骨已被完全吸收，胶原纤维逐渐降解，为萌出途径。在黏膜穿透途径中也发现了 Sema 3A 及其受体神经纤毛蛋白 1 和神经丛蛋白 A1 的类似阳性表达。Sema 3A 通过调节破骨细胞活性参与小鼠牙齿萌出途径的建立。Sema3A 可被视为小鼠牙齿萌出途径的新型神经递质或潜在生物标志物。
